| 1. |
- Gutierrez-Sanchez, Cristina, et al.
(författare)
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Gold nanoparticles as electronic bridges for laccase-based biocathodes
- 2012
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 134:41, s. 17212-17220
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Tidskriftsartikel (refereegranskat)abstract
- Direct electron transfer (DET) reactions between redox enzymes and electrodes can be maximized by oriented immobilization of the enzyme mols. onto an electroactive surface modified with functionalized Au nanoparticles (AuNPs). Here, the authors present such strategy for obtaining a DET-based laccase (Lc) cathode for O2 electroredn. at low overpotentials. The stable nanostructured enzymic electrode is based on the step-by-step covalent attachment of AuNPs and Lc mols. to porous graphite electrodes using the diazonium salt redn. strategy. Oriented immobilization of the enzyme mols. on adequately functionalized AuNPs allows establishing very fast DET with the electrode via their Cu T1 site. The measured electrocatalytic waves of O2 redn. can be deconvoluted into two contributions. The one at lower overpotentials corresponds to immobilized Lc mols. that are efficiently wired by the AuNPs with a heterogeneous electron transfer rate const. k0 » 400 s-1.
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| 2. |
- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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| 3. |
- Coman, Vasile, et al.
(författare)
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A membrane-, mediator-, cofactor-less glucose/oxygen biofuel cell.
- 2008
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Ingår i: Physical Chemistry Chemical Physics. - : Royal Society of Chemistry (RSC). - 1463-9084 .- 1463-9076. ; 10:40, s. 6093-6096
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Tidskriftsartikel (refereegranskat)abstract
- We report the fabrication and characterisation of a non-compartmentalised, mediator and cofactor free glucose-oxygen biofuel cell based on adsorbed enzymes exhibiting direct bioelectrocatalysis, viz. cellobiose dehydrogenase from Dichomera saubinetii and laccase from Trametes hirsuta as the anodic and cathodic bioelements, respectively, with the following characteristics: an open-circuit voltage of 0.73 V; a maximum power density of 5 muW cm(-2) at 0.5 V of the cell voltage and an estimated half-life of >38 h in air-saturated 0.1 M citrate-phosphate buffer, pH 4.5 containing 5 mM glucose.
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| 4. |
- Haberska, Karolina, et al.
(författare)
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Direct electron transfer reactions between human ceruloplasmin and electrodes.
- 2009
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Ingår i: Bioelectrochemistry. - : Elsevier BV. - 1878-562X .- 1567-5394. ; 76, s. 34-41
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Tidskriftsartikel (refereegranskat)abstract
- In an effort to find conditions favouring bioelectrocatalytic reduction of oxygen by surface-immobilised human ceruloplasmin (Cp), direct electron transfer (DET) reactions between Cp and an extended range of surfaces were considered. Exploiting advances in surface nanotechnology, bare and carbon-nanotube-modified spectrographic graphite electrodes as well as bare, thiol- and gold-nanoparticle-modified gold electrodes were considered, and ellipsometry provided clues as to the amount and form of adsorbed Cp. DET was studied under different conditions by cyclic voltammetry and chronoamperometry. Two Faradaic processes with midpoint potentials of about 400 mV and 700 mV vs. NHE, corresponding to the redox transformation of copper sites of Cp, were clearly observed. In spite of the significant amount of Cp adsorbed on the electrode surfaces, as well as the quite fast DET reactions between the redox enzyme and electrodes, bioelectrocatalytic reduction of oxygen by immobilised Cp was never registered. The bioelectrocatalytic inertness of this complex multi-functional redox enzyme interacting with a variety of surfaces might be associated with a very complex mechanism of intramolecular electron transfer involving a kinetic trapping behaviour.
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| 5. |
- Ressine, Anton, et al.
(författare)
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Bioelectrochemical studies of azurin and laccase confined in three-dimensional chips based on gold-modified nano-/microstructured silicon
- 2010
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Ingår i: Biosensors & Bioelectronics. - : Elsevier BV. - 1873-4235 .- 0956-5663. ; 25:5, s. 1001-1007
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Tidskriftsartikel (refereegranskat)abstract
- Double-sided three-dimensional porous silicon chips, 6 mm x 6 mm, covered with a 40 nm gold (nano)layer, were fabricated from a porous silicon wafer. Scanning electron microscopy along with electrochemical characterisation showed sample conductivity, mechanical stability, and high surface area of the thus fabricated devices, viz. 10 times higher electrochemically active surface area compared to the geometric area. The three-dimensional gold coated silicon chips were further modified with thiol layers, followed by immobilisation of a simple copper-containing redox protein, azurin, or a complex multicopper redox enzyme, laccase. The bioelectrochemical studies showed very high surface concentrations of azurin and laccase, i.e. close to the theoretical monolayer coverage. However, direct electron transfer reactions between the biomolecules and gold surfaces were observed only for a small percentage of the immobilised redox protein and enzyme, respectively. Thus, highly efficient oxygen-bioelectroreduction on laccase-modified 3D thiol-gold-porous silicon chips (as compared to planar laccase-modified gold electrodes, 42 mu A/cm(2) vs. 7 mu A/cm(2), respectively) was obtained only in the presence of an efficient soluble redox mediator. (C) 2009 Elsevier B.V. All rights reserved.
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| 6. |
- Vaz-Dominguez, Cristina, et al.
(författare)
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Combined ATR-SEIRAS and EC-STM study of the immobilization of laccase on chemically modified au electrodes
- 2012
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Ingår i: The Journal of Physical Chemistry C. - : American Chemical Society (ACS). - 1932-7447 .- 1932-7455. ; 116:31, s. 16532-16540
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Tidskriftsartikel (refereegranskat)abstract
- The successive steps of laccase immobilization on chem. modified Au electrodes were monitored using ATR-SEIRAS and in situ STM. Successful covalent immobilization of the enzyme on Au electrodes modified by a mixed aminophenyl-mercaptohexanol adlayer and on Au electrodes modified by a 4-aminothiophenyl SAM via a Schiff base reaction followed by the formation of amide bonds is revealed by the emergence of the corresponding bands in the ATR-SEIRA spectra, and an enzyme coverage on aminophenyl-mercaptohexanol-modified Au electrodes of about (7.27 ± 1.93) × 1011 laccase units per cm2 was calcd. from STM images. The small differences between the ATR-SEIRA spectra of the enzyme immobilized on aminophenyl-mercaptohexanol-modified Au electrodes and the ATR-SEIRA spectra of the enzyme immobilized on 4-aminothiophenyl-modified Au electrodes are attributed to a different orientation of the immobilized enzyme due to the presence on the surface of aminophenyl-mercaptohexanol-modified Au electrodes of OH functional groups that favor an orientation of laccase with the Cu T1 center of the enzyme facing the electrode surface, thus, allowing a high activity for direct electrocatalysis of the ORR at low overpotentials.
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| 7. |
- Vaz-Dominguez, Cristina, et al.
(författare)
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Laccase electrode for direct electrocatalytic reduction of O2 to H2O with high-operational stability and resistance to chloride inhibition
- 2008
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Ingår i: Biosensors & bioelectronics. - : Elsevier. - 0956-5663 .- 1873-4235. ; 24, s. 531-537
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Tidskriftsartikel (refereegranskat)abstract
- Laccase from Trametes hirsuta basidiomycete has been covalently bound to graphite electrodes electrochem. modified with Ph derivs. as a way to attach the enzyme mols. with an adequate orientation for direct electron transfer (DET). Current densities up to 0.5 mA/cm2 of electrocatalytic redn. of O2 to H2O were obtained in absence of redox mediators, suggesting preferential orientation of the T1 Cu center of the laccase towards the electrode. The covalent attachment of the laccase mols. to the functionalized electrodes permitted remarkable operational stability. Moreover, O2 bioelectroredn. based on DET between the laccase and the electrode was not inhibited by chloride ions, whereas mediated bioelectrocatalysis was. In contrast, fluoride ions inhibited both direct and mediated electron transfers-based bioelectrocatalytic redn. of O2. Thus, two different modes of laccase inhibition by halides are discussed.
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| 8. |
- Zumarraga, Miren, et al.
(författare)
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Combinatorial saturation mutagenesis of the Myceliophthora thermophila laccase T2 mutant : the connection between the C-terminal plug and the conserved 509VSG511 tripeptide
- 2008
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Ingår i: Combinatorial chemistry & high throughput screening. - : Bentham Science Publishers Ltd.. - 1386-2073 .- 1875-5402. ; 11:10, s. 807-816
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Tidskriftsartikel (refereegranskat)abstract
- A mutant laccase from the Ascomycete Myceliophthora thermophila has been submitted to iterative cycles of combinatorial satn. mutagenesis through in vivo overlap extension in Saccharomyces cerevisiae. Over 180,000 clones were explored, among which the S510G mutant revealed a direct interaction between the conserved 509VSG511 tripeptide, located in the neighborhood of the T1 site, and the C-terminal plug. The KmO2 value of the mutant increased 1.5-fold, and the electron transfer pathway between the reducing substrate and the T1 copper ion was altered, improving the catalytic efficiency towards non-phenolic and phenolic substrates by about 3- and 8-fold. Although the geometry at the T1 site was perturbed by the mutation, paradoxically the laccase redox potential was not significantly altered. Together, the results obtained in this study suggest that the 509VSG511 tripeptide may play a hitherto unrecognized role in regulating the traffic of oxygen through the C-terminal plug, the latter blocking access to the T2/T3 copper cluster in the native enzyme.
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